细胞色素P450花生四烯酸表氧化酶抑制TNF-α诱导的内皮细胞凋亡  

Arachidonic acid cytochrome P450 epoxygenases protect vessel endothelial cells against apoptosis induced by tumor necrosis factor-α

作　　者：林立[1] 王红[1] 王炎[1] 王燕[1] 陆再英 汪道文[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院心血管研究室暨基因治疗中心,湖北武汉430030

出　　处：《中国病理生理杂志》2007年第10期1878-1882,共5页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.39870307)

摘　　要：目的:以TNF-α/act D诱导的细胞凋亡作为一种内皮细胞损伤的模型,通过转染细胞色素P450表氧化酶基因BM3.F87V、2C11OR以及2J2产生内源性EDHF(EETs),探讨EDHF是否具有抗内皮细胞损伤的作用及其机制。方法:以CYP450表氧化酶BM3.F87V、2C11OR及2J2基因转染3-4代培养的牛主动脉血管内皮细胞。通过MTT方法检测细胞活性。基因转染24 h后,用TNF-α/act D作用一定时间诱导内皮细胞凋亡,以细胞形态学观察,DNA ladder分析,流式细胞仪分析(Annexin-V-FITC和propidium iodide双染法)作为检测细胞凋亡的指标。结果:MTT分析显示,转染CYP450 BM3.F87V、2C11OR以及CYP2J2基因的内皮细胞活性(分别为189.0%±18.0%,166.0%±22.6%或131.0%±22.8%)强于对照组(100.0%±21.3%)。经TNF-α/act D诱导,转染pCB6的对照组内皮细胞大量脱落,未脱落细胞中许多细胞核呈碎裂颗粒状(约20.1%±0.9%);而转染CYPBM3.F87V、CYP2C11OR或CYP2J2基因的内皮细胞仅见少数核呈碎颗粒状(分别约1.2%±0.2%,1.0%±0.2%或3.1%±0.2%)。DNA ladder分析显示TNF-α/act D诱导后,转染表氧化酶基因的内皮细胞所形成的梯形条带弱于对照组。进一步用流式细胞仪定量检测表明,转染细胞色素表氧化酶基因CYP BM3F87V或2C11OR内皮细胞中凋亡细胞所占比例(分别为12.35%±4.55%,10.92%±3.57%)明显低于对照组(29.57%±2.84%),而转染CYP2J2的细胞凋亡率也明显低于对照组。结论:转染CYP450基因能够增加内皮细胞活性,抑制TNF-α诱导的内皮细胞凋亡,具有内皮细胞保护效应。AIM： To investigate whether endogenous endothelium -derived hyperpelarizing factors （EHDFs） produced by CYP epoxygenases BM3 · F87V, 2CllOR or CYP2J2 transfection was able to protect endothelial cells against apoptosis induced by tumor necrosis factor alpha. METHODS ： Three or four passages of cultured bovine aortic endothelial cells （BAECs） were transfected with epoxygenases or the empty vector （ pCB6 ）. Cell viability was detected by MTT assay. Apoptosis of transfected endothelial cells was evaluated by DNA ladder assay, flow cytometry and morphological observations under fluorescence microscopy. RESULTS： Overexpression of CYP epoxygenases BM3· F87V, 2CllOR, CYP2J2 increased cell viability respectively observed by MTT assay. The percentage of cells undergoing apeptosis was decreased in 2C11OR-, BM3F87V- or 2J2 -transfected cells compared to the vector as evaluated by DNA fragment assay, flow cytometry analysis and morphological observations under fluorescence microscopy. CONCLUSION： Overexpression of CYP epexygenases BM3 · F87V, 2 C11 OR or 2.J2 increases cell viability and protects endothelial cells against TNF -α - induced apeptosis. These findings suggest new targets to investigate the endothelium - associated disorders and provide novel thera-peutic strategies to treat them by modulating cytochrome P450 epoxygenases.

关 键 词：内皮源性超极化因子 内皮细胞 环氧二十碳烯酸 细胞凋亡 

分 类 号：R363[医药卫生—病理学]